Nucleic acid molecules that target RPS6 and confer resistance to coleopteran pests

Abstract

This disclosure concerns nucleic acid molecules and methods of use thereof for control of coleopteran pests through RNA interference-mediated inhibition of target coding and transcribed non-coding sequences in coleopteran pests. The disclosure also concerns methods for making transgenic plants that express nucleic acid molecules useful for the control of coleopteran pests, and the plant cells and plants obtained thereby.

What is claimed is: 
     
       1. A transgenic plant cell comprising a polynucleotide comprising at least one nucleotide sequence(s) selected from the group consisting of:
 SEQ ID NO:7; 
 the complement of SEQ ID NO:7 
 a native coding sequence comprising SEQ ID NO:7 of a  Diabrotica  organism; 
 the complement of a native coding sequence comprising SEQ ID NO:7 of a  Diabrotica  organism; 
 SEQ ID NO:28; and 
 the complement of SEQ ID NO:28, 
 wherein the polynucleotide is expressed in the transgenic plant cell as a ribonucleic acid molecule, and wherein contacting the ribonucleic acid molecule with a coleopteran pest inhibits the expression of an endogenous nucleic acid comprising a nucleotide sequence specifically complementary to the nucleotide sequence comprised in the polynucleotide. 
 
     
     
       2. The transgenic plant cell of  claim 1 , wherein the at least one nucleotide sequence(s) is operably linked to a heterologous promoter. 
     
     
       3. The transgenic plant cell of  claim 1 , wherein the  Diabrotica  organism is selected from the group consisting of  Diabrotica virgifera virgifera  LeConte;  Diabrotica barberi  Smith and Lawrence;  Diabrotica undecimpunctata howardi; Diabrotica virgifera zeae; Diabrotica balteata  LeConte;  Diabrotica undecimpunctata tenella ; and  Diabrotica undecimpunctata undecimpunctata  Mannerheim. 
     
     
       4. The transgenic plant cell of  claim 1 , wherein the polynucleotide further comprises at least one nucleotide sequence encoding a polypeptide from  Bacillus thuringiensis.    
     
     
       5. The transgenic plant cell of  claim 4 , wherein the polypeptide from  B. thuringiensis  is selected from the group consisting of Cry3, Cry34, and Cry35. 
     
     
       6. The transgenic plant cell of  claim 1 , wherein the polynucleotide is transcribed to produce a double-stranded ribonucleic acid molecule in the cell. 
     
     
       7. The transgenic plant cell of  claim 6 , wherein contacting the double-stranded ribonucleic acid molecule with the coleopteran pest kills or inhibits the growth, reproduction, and/or feeding of the coleopteran pest. 
     
     
       8. The transgenic plant cell of  claim 6 , wherein the double-stranded ribonucleic acid molecule comprises a first, a second, and a third polyribonucleotide,
 wherein the first polyribonucleotide comprises the polyribonucleotide encoded by the at least one nucleotide sequence(s), 
 wherein the third polyribonucleotide is linked to the first polyribonucleotide by the second polyribonucleotide, and 
 wherein the third polyribonucleotide is substantially the reverse complement of the first polyribonucleotide, such that the portions of the ribonucleic acid molecule comprising each of the first and the third polyribonucleotides hybridize to each other in the double-stranded ribonucleotide molecule. 
 
     
     
       9. The transgenic plant cell of  claim 1 , wherein the polynucleotide is operably linked to a heterologous promoter functional in a plant cell. 
     
     
       10. A plant comprising the transgenic plant cell of  claim 9 . 
     
     
       11. A seed of the plant of  claim 10 , wherein the seed comprises the polynucleotide. 
     
     
       12. The plant of  claim 10 , wherein the polynucleotide is expressed in the plant as a double-stranded ribonucleic acid molecule. 
     
     
       13. The plant of  claim 10 , wherein the plant is  Zea mays.    
     
     
       14. The plant of  claim 10 , wherein the polynucleotide is expressed in the plant as a ribonucleic acid molecule, and the ribonucleic acid molecule inhibits the expression of an endogenous coleopteran pest nucleotide sequence specifically complementary to the at least one nucleotide sequence(s) when the coleopteran pest ingests a part of the plant. 
     
     
       15. A commodity product produced from a plant according to  claim 10 , wherein the commodity product comprises a detectable amount of the polynucleotide. 
     
     
       16. The transgenic plant cell of  claim 9 , wherein the cell is a  Zea mays  cell. 
     
     
       17. A method for improving the yield of a corn crop, the method comprising cultivating a transgenic corn plant comprising the transgenic corn cell of  claim 16  to allow the expression of a nucleic acid molecule comprising the polyribonucleotide encoded by the at least one nucleotide sequence(s), wherein expression of the nucleic acid molecule inhibits coleopteran pest infection or growth and loss of yield due to coleopteran pest infection. 
     
     
       18. The method according to  claim 17 , wherein the nucleic acid molecule suppresses at least a first target gene in a coleopteran pest that has contacted a portion of the corn plant. 
     
     
       19. A method for controlling a coleopteran pest population, the method comprising cultivating a host plant of a coleopteran pest,
 wherein the host plant comprises the transgenic plant cell of  claim 9 , and 
 wherein the polynucleotide is expressed to produce a ribonucleic acid molecule that functions upon contact with a coleopteran pest belonging to the population to inhibit the expression of a target sequence within the coleopteran pest and results in decreased growth of the coleopteran pest or coleopteran pest population, relative to growth on a host plant of the same species lacking the transformed plant cell. 
 
     
     
       20. The method according to  claim 19 , wherein the ribonucleic acid molecule is a double-stranded ribonucleic acid molecule. 
     
     
       21. The method according to  claim 19 , wherein the coleopteran pest population is reduced relative to a coleopteran pest population infesting a host plant of the same species lacking the transformed plant cell. 
     
     
       22. A method of controlling plant coleopteran pest infestation in a plant, the method comprising providing in the diet of a coleopteran pest the transgenic plant cell of  claim 9 . 
     
     
       23. A method for selecting a transgenic plant cell, the method comprising:
 culturing the transgenic plant cell of  claim 9  under conditions sufficient to allow for development of a plant cell culture comprising a plurality of transgenic plant cells; 
 selecting for transgenic plant cells that have integrated the polynucleotide into their genomes; 
 screening the transgenic plant cells for expression of a ribonucleic acid molecule comprising the polyribonucleotide encoded by the at least one nucleotide sequence(s); and 
 selecting a transgenic plant cell that expresses the ribonucleic acid molecule. 
 
     
     
       24. A method for producing a coleopteran pest-resistant transgenic plant, the method comprising regenerating a transgenic plant from the transgenic plant cell of  claim 1 , wherein expression of the ribonucleic acid molecule encoded by the polynucleotide is sufficient to inhibit the expression of the endogenous nucleic acid when the coleopteran pest feeds on the transgenic plant. 
     
     
       25. The transgenic plant cell of  claim 1 , wherein the polynucleotide is SEQ ID NO:31.